Figure 6

Myogenin from myotube nuclear proteins extracts binds to the wild-type ART1 promoter E box. Gelshift assays were performed by incubation of the [α-³²P]dCTP labelled probe containing the E box (WT) of the ART1 gene promoter with nuclear protein extracts (NE) of C3H-10T 1/2 (MyoD-Mb, Mt) and C2C12 (Mb, Mt) cells. The specificity of the protein binding at the E box site was assessed by the use of a labelled probe containing a mutated ART1 E box sequence (Mut). For supershift assays, reaction mixtures were preincubated with antibodies (Ab) against myogenin (αMG) or with a non-specific isotype (ISO) as a negative control. Specific retarded protein/probe complexes are marked by white arrows and supershifted complexes by black arrows. As a positive control a probe containing a myogenin binding E box of the desmin gene promoter [43] was used.